ObjectiveTo investigate the inhibitory effect of peroxisome proliferator-activated receptor-?(PPAR?) agonist on mesangial cell(MC) proliferation and extracellular matrix expression induced by angiotensin Ⅱ(Ang Ⅱ).MethodsThe incorporation of 3H-thymidine(3H-TdR) and cell count were used as the measurement of MC proliferation.MC cell-cycle was analyzed by flow cytometry.Mouse primary MC was treated with various concentration of Ang Ⅱ(1,10,100 nmol/L) in the presence or Absence of N-acytosistin(NAC) or rosiglitazone.Transforming growth factor-?1(TGF-?1),plasminogen activator inhibitor-1(PAI-1),and fibronectin(FN) mRNA expression were determined by real time-PCR.Reactive oxygen species(ROS) production was measured by 2,7-dichlorofluorescein diacetate(DCFDA) fluorescence.Results1.One hundred nmol/L Ang Ⅱ increased 3H-TdR incorporation and cell number by 2.14 and 2.32 fold,respectively.Ang Ⅱ-induced MC proliferation was inhibited by PPAR? agonist rosiglitazone with dose-dependent manner in mouse MC.2.One hundred nmol/L Ang Ⅱ stimulation for 24 h induced 48% MC processed to S and G2/M phase.Rosiglitazone significantly blocked Ang Ⅱ increased cell number in S and G2/M phase.3.Rosiglitazone reduced Ang Ⅱ-induced TGF-?1,PAI-1,and FN mRNA expression with dose-dependent manner.4.One hundred nmol/L Ang Ⅱ stimulation for 60 min increased ROS production by 3.85 folds.Rosiglitazone significantly inhibited Ang Ⅱ-induced ROS production.Ten ?mol/L rosiglitazone almost completely blocked Ang Ⅱ-induced ROS production.ConclusionPPAR? agonist rosiglitazone could block Ang Ⅱ-induced MC proliferation and extracellular matrix expression via inhibition of ROS production.

Adult ; Aged ; Female ; Humans ; Liver Transplantation ; adverse effects ; Male ; Middle Aged ; Postoperative Hemorrhage ; etiology ; Tracheotomy ; adverse effects ; Young Adult

Cyclohexanones ; poisoning ; Ethylene Dichlorides ; poisoning ; Humans ; Infant ; Male

Objective: To chine and express the recombinant human endothelial monocyte-activating polypeptide-Ⅱ(EMAP-Ⅱ)and identify its anti-tumor biological activities.Methods: EMAP-Ⅱ_(147-312)was expressed by the expression vector pMAL-p2x and E.coli BL-21 and the product was purified.The production of tissue factor(TF)in human umbili- cal vein endothelial cell ECV-304 mediated by the recombinant EMAP-Ⅱwas determined by chemiluminescence sub- strate.The promoting effect of recombinant EMAP-Ⅱon TNF?-induced ECV-304 cell.Apoptosis was determined by flow cytometry.Its inhibitory effect on human pancreaic cancer cell SW1990 proliferation was determined by MTT method. Results:DNA sequencing verified that EMAP-Ⅱwas correctly cloned.The molecular mass of the protein identified by SDS-PAGE was consistent with the theoretic value.The productivity of recombinant EMAP-Ⅱwas 500?g per 1 g bacteria (wet mass).The purified product induced expression of tissue factor(TF)in ECV-304 cells;it also enhanced the sensi- tivity of ECV-304 cells to the apoptotic effect of TNF?([16.6?2.5]% vs[25.6?2.3]%,P

OBJECTIVETo construct the prokaryotic expression systems of tpnl7 and tpn47 genes and tpn17-tpn47 fusion of Treponema pallidum, and to establish ELISAs based on rTpN17, rTpN47 and rTpN17-TpN47 as antigens to evaluate the sensitivity and specificity of the ELISAs for detection of serological diagnosed syphilis.

METHODStpn17 and tpn47 genes were amplified and cloned by routine molecular biological methods. PCR with linking primers was used to construct artificial fusion gene tpn17-tpn47. The prokaryotic expression systems of the genes were then constructed. SDS-PAGE was used to measure the expression of the target recombinant proteins rTpN17, rTpN47 and rTpN17-TpN47. Ni-NTA affinity chromatography was applied to extract the three recombinant proteins, while Western blot was performed to determine their immunity. Using rTpN17, rTpN47 and rTpN17-TpN47 as the coated antigens, ELISAs (rTpN17-ELISA, rTpN47-ELISA and rTpN17-TpN47- ELISA) were established to detect serum samples from 200 healthy individuals, 25 RA patients, 17 SLE patients and 419 syphilis patients. Results of the ELISAs were compared to those with TRUST and TPHA.

RESULTSThe sequence similarities of the cloned tpnl7 and tpn47 genes and the constructed tpn17-tpn47 fusion gene were 100%, compared with the corresponding sequences in GenBank. The expression outputs of rTpN17, rTpN47 and rTpN17-TpN47 were 37.2%, 23.3% and 29.8% of the total bacterial proteins, respectively. Each of the three purified recombinant proteins showed a single fragment in gel after electrophoresis, and could take place remarkable conjugation reactions to the positive sera from syphilis patients. The detection results of rTpN17-ELISA, rTpN47-ELISA and rTpN17-TpN47-ELISA were negative for the serum samples from healthy individuals, RA and SLE patients, while presented 84.4%, 82.3% and 98.1% positive detection rates for the serum samples from syphilis patients. The positive detection rates of rTpN17-ELISA and rTpN47-ELISA were lower than that of TPHA (P<0.01), while the positive detection rate of rTpN17-TpN47-ELISA was similar to that of TPHA (P>0.05). All the positive detection rates from ELISA tests were higher than that of TRUST (71.4%).

CONCLUSIONrTpN17-ELISA, rTpN47-ELISA and especially rTpN17-TpN47-ELISA established in this study were of great hope as it was rapid, simple, convenient, safe, with high sensitivity and specificity for serological screening and detection of syphilis.

Bacterial Proteins ; biosynthesis ; genetics ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; methods ; Genes, Bacterial ; Humans ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Sensitivity and Specificity ; Syphilis ; diagnosis ; Syphilis Serodiagnosis ; methods ; Treponema pallidum ; genetics

OBJECTIVETo construct a prokaryotic expression system to serially express Alloferon-1 and to determine the anti-tumor activity of its products in vitro.

METHODSAn artificial fusion gene containing 6 x His-EK-8 x Alloferon-1-EK-6 x His sequences was constructed by linking primer PCR. By using routine molecular biological methods, the artificial fusion gene was cloned and its prokaryotic expression system was then constructed. SDS-PAGE and BioRad Agarose Image Analysor was applied to measure the expression and output of the target recombinant products 8 x rAlloferon-1-EK. Ni-NTA affinity chromatography and EK digestion and Sephadex G-50 chromatography were performed to extract 8 x rAlloferon-1-EK and rAlloferon-1-EK, respectively. The proliferation of KB, SGC and HL-60 tumor cells was tested by using MTT method after treatment with directly synthesized Alloferon-1 (sAlloferon-1), Aloferon-1-EK (sAlloferon-1-EK) and rAlloferon-1-EK.

RESULTThe target artificial fusion gene and its prokaryotic expression system pET42a-8 x rAlloferon-1-EK-E. coliBLDE3 with the expected sequences were obtained. Under inducement of IPTG, the prokaryotic expression system expressed the target serial recombinant protein 8 x rAlloferon-1-EK and its output was approximate 30 % of the total bacterial proteins. 8 x rAlloferon-1-EK and rAlloferon-1-EK were obtained through Ni-NTA and Sephadex G-50 columns. sAlloferon-1, sAlloferon-1-EK and rAlloferon-1ìrAlloferon-EK showed similar remarkable effects of inhibiting the growth and proliferation of KB, SGC and HL-60 cells in vitro within 25 approximately 100 microg/ml concentration range (P<0.01), and there were no significant differences in the inhibiting effects among the three agents (P>0.05).

CONCLUSIONA prokaryotic expression system to serially express rAlloferon-1 has been successfully constructed. The product rAlloferon-1-EK has a similar anti-tumor activity compared to both the synthesized Alloferon-1 and Alloferon-1-EK in vitro.

Animals ; Anti-Bacterial Agents ; pharmacology ; Antineoplastic Agents ; metabolism ; pharmacology ; Bacterial Vaccines ; administration & dosage ; genetics ; metabolism ; Cell Line, Tumor ; Escherichia coli ; genetics ; metabolism ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Growth Inhibitors ; genetics ; metabolism ; pharmacology ; HL-60 Cells ; Helicobacter pylori ; genetics ; metabolism ; Humans ; KB Cells ; Peptides ; genetics ; immunology ; metabolism ; pharmacology ; Recombinant Fusion Proteins ; genetics ; metabolism ; pharmacology

PURPOSE: The purposes of this study were to examine the level of social support, health promoting behaviors and depression among unmarried pregnant women and to identify the relationship between social support, health promoting behaviors and depression. METHOD: A descriptive correlational study was conducted. The participants were 102 unmarried pregnant women receiving shelter services from four facilities in two metropolitan cities. Data was collected using a self-administered questionnaire. Descriptive statistics, ANOVA and Pearson correlation were used for data analysis. RESULTS: The level of social support and health promoting behaviors were relatively lower and the level of depression was relatively higher than those of married pregnant women. The participants received especially low social support from their unmarried partner. There was a positive relationship between social support and health promoting behaviors. Moreover, there were negative relationships between social support and depression and between health promoting behaviors and depression. CONCLUSIONS: To promote physical and emotional health of unmarried pregnant women, more attention is necessary to increase their social support. A nursing intervention program to increase social support among unmarried pregnant women in needed.
Depression* ; Female ; Humans ; Nursing ; Pregnant Women* ; Surveys and Questionnaires ; Single Person* ; Statistics as Topic

Female ; Humans ; Laser Therapy ; methods ; Middle Aged ; Semicircular Canals ; surgery ; Vertigo ; etiology ; surgery

Background To suppress vascular endothelial growth factor (VEGF) is a researching hot topic for the treatment and prevention of retinal neovascularization.Some detectable efficacy of VEGF small interference RNA (VEGF siRNA) in anti-tumor neovascularization has been well-known.But relevant study on VEGF siRNA on retinal neovascularization is seldom.Objective Present study was to investigate the inhibiting effect of VEGF siRNA on retinal neovascularization.Methods The 48 clean C57BL/6J mice aged 7-day-old were randomly divided into normoxia group,hypoxia control group,vector group and VEGF siRNA group and 12 mice for each.Hypoxia models were established by raising the pups with mother mice in the airtight oxygen-cabin for 5 days.The lipofectamineTM 2000 (LF2000)-mediated vector plasmids or VEGF siRNA recombinant plasmids were then injected intravitreally in 12 12-day-old pup mice respectively.The animals were sacrificed in 1 week after intravitreal injection,and the numbers of vascular endothelial cell nuclei extending beyond the internal limiting membrane (ILM) were counted by hematoxylin-eosin stain.The expressions of VEGF protein and mRNA in retina were assayed by immunoinfluorescence technique and RT-PCR.Results The numbers of vascular endothelial cell nuclei extending beyond the ILM were 0.19±0.09,24.89±2.03,23.65±2.15 and 8.83±1.12 in normoxia group,model control group,vector group and VEGF siRNA group separately,showing significant decrease in VEGF siRNA group compared with model control group or vector group (q=5.67,q=4.97,P＜0.01).RT-PCR revealed that VEGF mRNA was faintly expressed in mouse retina in normoxia group.However,in model control group and vector group,the level of VEGF mRNA was 52.3 times and 36.7 times more than that of normoxia group respectively and only 3.5 times in VEGF siRNA group,presenting a inhibitory rate of 43.39% of VEGF siRNA on VEGF.Immunofluorescence showed that the expression of VEGF was weaker in normoxia group and strong positive response in model control group and vector group,but the expression intensity of VEGF protein was significantly weaker in VEGF siRNA group.Conclusion VEGF siRNA recombinant plasmids can efficiently inhibit retinal neovascularization in oxygen-induced retinopathy mouse model through intravitreal injection.

Objective To study the changes of serum interleukin(IL)-4,IL-12 and correlation with cellular immunity in children with asthma of different stages.Methods Fifty asthmatic children were randomly selected, including 30 cases in attack stage (group A) and 20 cases in remission stage (group R). At the same time, 22 healthy children were studied as normal controls (group N).The levels of IL-12 and IL-4 ,T cells subgroups and erythrocyte immunity were detected.Results 1.Serum IL-12 levels were (24.44? 13.26 ),(42.30?12.65),(44.68?28.28) ng/L in group A, R and N,respectively. There was significant difference in three groups (F=8.92 P